CD4 is a cell surface glycoprotein of CD4.sup.+ T lymphocytes (helper/inducer cells). CD4.sup.+ lymphocytes are critical regulatory cells of the human immune system. They mediate T cell proliferation, lymphokine release and helper cell interactions affecting immunoglobulin release. The primary targets of certain infective agents, including the human immunodeficiency virus ("HIV"), are cells bearing the CD4 glycoprotein.* FNT * This application uses the generic term human immunodeficiency virus ("HIV") to refer to independent isolates from AIDS patients and to laboratory strains derived therefrom. The term HIV includes viruses elsewhere identified as human T cell lymphotrophic virus type III ("HTLV-III"), lymphadenopathy-associated virus ("LAV") and AIDS-associated retrovirus ("ARV"). The HIV terminology was adopted by the human retrovirus subcommittee of the International Committee On Taxonomy Of Viruses.
Such cells include CD4.sup.+ lymphocytes, macrophages and certain brain cells.
Upon infection with HIV, CD4.sup.+ lymphocytes are rendered non-functional and become depleted. This T cell depletion has been attributed both to recurrent cycles of infection resulting from lysis of infected cells and to fusion (syncytia formation) between CD4.sup.+ infected and uninfected cells (J. Sodroski et al., "Role of The HTLV-III/LAV Envelope In Syncytium Formation And Cytopathicity", Nature, 322, pp. 470-74 (1986)3. The depletion of CD4.sup.+ lymphocytes leads to immunosuppression, with the patient becoming susceptible to a wide range of opportunistic infections and malignancies. Such immunosuppression is seen in patients suffering from acquired immunodeficiency syndrome ("AIDS"). In some cases, AIDS is accompanied by central nervous system disorders, thought to be directly caused by HIV infection of CD4.sup.+ brain cells. Complete clinical manifestation of AIDS is usually preceded by AIDS related complex ("ARC"), a syndrome characterized by symptoms such as persistent generalized lymphadenopathy, fever and weight loss. HIV is thought to be the etiological agent for AIDS and its precursor, ARC M. G. Sangadharan et al., "Detection, Isolation And Continuous Production Of Cytopathic Retroviruses (HTLV-III) From Patients With AIDS And Pre-AIDS", Science, 224, pp. 497-508 (1984)!.
The major surface (envelope) protein of HIV is produced as a precursor polypeptide (gp160) which, in mature form, is cleaved into a large heavily glycosylated exterior membrane protein having about 481 amino acids--gp120--and a smaller transmembrane protein of about 345 amino acids, which may be glycosylated--gp41 L. Ratner et al., "Complete Nucleotide Sequence Of The AIDS Virus", HTLV-III, Nature, 313, pp. 277-84 (1985)!.
It is believed that HIV gp120 selectively binds to CD4 epitope(s), thus targeting HIV to CD4.sup.+ cells A. G. Dalgleish et al., "The CD4 (T4) Antigen Is An Essential Component Of The Receptor For The AIDS Retrovirus", Nature, 312, pp. 763-67 (1984); D. Klatzmann et al., "T-Lymphocyte T4 Molecule Behaves As The Receptor For Human Retrovirus LAV", Nature, 312, pp. 767-68 (1984)!. The binding of HIV gp120 to cell surface CD4 is thought to initiate infection of the CD4.sup.+ cell. It is also thought to initiate membrane fusion of infected CD4.sup.+ cells with uninfected CD4.sup.+ cells (i.e., syncytia formation), which contributes to cell-to-cell transmission of the virus and to its cytopathic effects J. A. Habeshaw and A. G. Dalgleish, "The Relevance Of HIV env/CD4 Interactions To The Pathogenesis Of Acquired Immune Deficiency Syndrome", J. AIDS, 2, pp. 457-68 (1989); J. D. Lifson and E. G. Engleman, "Role Of CD4 In Normal Immunity And HIV Infection", Immunol. Rev., 109, pp. 93-117 (1989)!.
Mature CD4 (also known as T4) is a 433 amino acid glycoprotein displaying a molecular mass of 55,000 to 62,000 daltons, having an extracellular domain (approximately AA.sub.1 -AA.sub.375), a membrane spanning domain (approximately AA.sub.376 -AA.sub.395) and a cytoplasmic tail (approximately (AA.sub.396 -AA.sub.433). CD4 is synthesized as a pre-protein with a 25 amino acid signal sequence. The nucleotide sequence and deduced amino acid sequence for cDNA encoding full length human CD4 have been reported P. J. Maddon et al., "The Isolation And Nucleotide Sequence Of A cDNA Encoding The T cell Surface Protein T4: A New Member Of The Immunoglobulin Gene Family", Cell, 42, pp. 93-104 (1985); D. R. Littman et al., "Corrected CD4 Sequence", Cell, 55, p. 541 (1988)!. The CD4 extracellular domain consists of four tandem regions having homology to immunoglobulin V regions--V1 (spanning approximately AA.sub.1 -AA.sub.100), V2 (spanning approximately AA.sub.101 -AA.sub.180), V3 (spanning approximately AA.sub.181 -AA.sub.290) and V4 (spanning approximately AA.sub.291 -AA.sub.375 see Maddon et al., Cell, supra; J. Wang et al., "Atomic Structure Of A Fragment Of Human CD4 Containing Two Immunoglobulin-Like Domains", Nature, 348, pp. 411-18 (1990)!.
The CD4 V1 region has been identified as the binding site of HIV gp120 J. Arthos et al., "Identification Of The Residues In Human CD4 Critical For The Binding Of HIV", Cell, 57, pp. 469-81 (1989); T. Mizukami et al., "Binding Region For Human Immunodeficiency Virus (HIV) And Epitopes For HIV-Blocking Monoclonal Antibodies Of The CD4 Molecule Defined By Site-Directed Mutagenesis", Proc. Natl. Acad. Sci. USA, 85, pp. 9273-77 (1988); A. Peterson and B. Seed, "Genetic Analysis Of Monoclonal Antibody And HIV Binding Sites On The Human Lymphocyte Antigen CD4", Cell, 54, pp. 65-72 (1988); N. R. Landau et al., "The Envelope Glycoprotein Of The Human Immunodeficiency Virus Binds To The Immunoglobulin-Like Domain Of CD4", Nature, 334, pp. 159-62 (1988); L. K. Clayton et al., "Substitution Of Murine For Human CD4 Residues Identifies Amino Acids Critical For HIV-gp120 Binding", Nature, 335, pp. 363-66 (1988)!.
The possible ability of anti-CD4 antibodies to prevent or treat HIV infection by blocking HIV gp120 binding to CD4 has led a number of workers to explore the ability of certain anti-CD4 antibodies to block HIV-induced syncytia formation between CD4.sup.+ cells, as well to study the immunosuppressiveness of some of those antibodies.
For example, the CD4 V1-specific antibodies Leu3A and OKT4A have been reported to be effective blockers of HIV-induced syncytia formation Q. J. Sattentau, "Epitopes Of The CD4 Antigen And HIV Infection", Science, 234, pp. 1120-23 (1986); B. A. Jameson et al., "Location And Chemical Synthesis Of A Binding Site For HIV-1 On The CD4 Protein", Science, 240, pp. 1335-39 (1988); Peterson and Seed, Cell, supra!. However, these antibodies have serious drawbacks for use as a pharmaceutical for treatment of HIV infection. For example, they can not bind to or act on a CD4 molecule which is already bound to HIV gp120 , because OKT4A and Leu3A recognize CD4 epitopes that overlap with the gp120 binding site see, e.g., P. A. Bates et al., "A Predicted Three-Dimensional Structure For The Human Immunodeficiency Virus Binding Domains Of CD4 Antigen", Prot. Engng., 3, pp. 13-21 (1989); J. S. McDougal et al., "Binding Of The Human Retrovirus HTLV-III/LAV/ARV/HIV To The CD4 (T4) Molecule: Conformation Dependence, Epitope Mapping, Antibody Inhibition, And Potential For Idiotypic Mimicry", J. Immunol., 137, pp. 2937-44 (1986); Landau et al., Nature, supra; A. G. Dalgleish et al., "Neutralisation Of HIV Isolates By Anti-Idiotypic Antibodies Which Mimic The T4 (CD4) Epitope: A Potential AIDS Vaccine", Lancet, 2, pp. 1047-50 (1987)!.
Also, OKT4A has been reported to be quite immunosuppressive, which is not a desirable characteristic for a putative AIDS therapeutic D. Lamarre et al., "Class II MHC Molecules And The HIV gp120 Envelope Protein Interact With Functionally Distinct Regions Of The CD4 Molecule", EMBO J., 8, pp. 3271-77 (1989); W. E. Biddison et al., "Possible Involvement Of The OKT4 Molecule In T Cell Recognition Of Class II HLA Antigens", J. Exp. Med., 156, pp. 1065-76 (1982)!.
Anti-CD4 monoclonal antibodies that are specific for other CD4 epitopes or domains have been studied, but the published reports on these antibodies also evidence drawbacks for use as AIDS therapeutics.
One such antibody is OKT4B, which has been reported to be specific for the V2 domain of CD4 T. Kieber-Emmons et al., "The gp120-CD4 Interface: Structural, Immunological And Pathological Considerations", Biochim. Biophys. Acta, 989, pp. 281-300 (1989)!. There have been conflicting reports as to whether OKT4B significantly interferes with HIV gp120 binding to CD4 McDougal et al., J. Immunol., supra; K. Lundin et al., "A Specific Assay Measuring Binding Of .sup.125 I-GP120 From HIV To T4.sup.+ /CD4.sup.+ Cells", J. Immunol. Methods, 97, pp. 93-100 (1987); Lamarre et al., EMBO J., supra!. And, OKT4B has been reported to be significantly more immunosuppressive than OKT4A Lamarre et al., EMBO J., supra!. Moreover, OKT4B is a relatively weak blocker of HIV-induced syncytia formation, as compared to OKT4A Sattentau et al., Science, supra!.
Other anti-CD4 antibodies that have been reported to have some effect on HIV-induced syncytia formation, and also have been reported to bind to CD4 epitopes distinct from the epitopes bound by OKT4A and Leu3A, include MT151, VIT4 and MT321 Sattentau et al., Science, supra!. However, several more recent independent studies indicate that these closely related antibodies recognize conformational-dependent CD4 epitopes that overlap with the CD4 epitope involved in gp120 binding Q. J. Sattentau et al., "Structural Analysis Of The Human Immunodeficiency Virus-Binding Domain Of CD4", J. Exp. Med., 170, pp. 1319-34 (1989); Bates et al., Prot. Engng., supra; Landau et al., Nature, supra, M. Merkenschlager et al., "Functional Epitope Analysis Of The Human CD4 Molecule", J. Immunol., 9, pp. 2839-45 (1990)!.
Another anti-CD4 antibody that has been studied is OKT4, which is specific for the V3V4 domain of CD4 E. A. Berger et al., "A Soluble Recombinant Polypeptide Comprising The Amino-Terminal Half Of The Extracellular Region Of The CD4 Molecule Contains An Active Binding Site For Human Immunodeficiency Virus", Proc. Natl. Acad. Sci. USA, 85, pp. 2357-61 (1988)!. OKT4 has even greater drawbacks as a therapeutic agent for HIV infection than OKT4A, Leu3A, OKT4B and other known anti-CD4 antibodies. For example, OKT4 has been reported to be a "nonblocker" of HIV-induced syncytia formation Sattentau, Science, supra!.
Before our invention, no anti-CD4 antibody had been reported to be capable of both not significantly blocking the binding of HIV gp120 to human CD4 and effectively blocking HIV-induced syncytia formation. Such an antibody would clearly offer advantages for therapeutic intervention in AIDS, ARC and HIV infection, as it could be used to intervene after HIV binding to CD4, or in combination with agents that do block the HIV gp120-CD4 binding event. Thus, the need exists for anti-CD4 antibodies having such a highly desirable combination of properties, for use in the treatment and prevention of AIDS, ARC and HIV infection.